Epstein-Barr virus is a human herpesvirus that can latently infect and immortalize human B lymphocytes in vitro. The virus persists for life in vivo and can give rise to lymphoma in immunosuppressed individuals. Although infection and latency have been studied extensively in vitro, little is known about latent infection in vivo. The aim of this proposal is to characterize latent infection by Epstein-Barr virus in vivo and how that latency is perturbed by immunosuppression. We have developed a highly sensitive DNA PCR assay for EBV which allows us to detect a single genome in as many as 5x10/7 uninfected cells. We have used this assay to measure the frequency of virus infected cells in the peripheral blood of healthy individuals and to demonstrate that the phenotype of these cells is uniquely different (CD23- B cells) from that of in vitro, EBV infected lymphoblasts. We intend to further characterize the cell surface phenotype of the cells (activated vs. resting, memory vs. virgin, mature vs. BL/germinal center like, Bi vs. B2). This information will identify target cell(s) for EBV latency in vivo and be used to develop protocols for enriching cell populations that carry the virus, which can then be tested for viral gene expression using RNA PCR. We hypothesize that the infected cells in healthy individuals will be resting and expressing a limited set of latent genes, so as to avoid immunosurveillance. Upon immunosuppression the frequency of these cells should not change, however, cells resembling typical EBV lymphoblasts, normally under immunosurveillance and potential precursors for immunoblastic lymphoma, will now be detected. These studies will be carried out in collaboration with the transplantation unit at the New England Medical Center with the long term goal of identifying potential risk factors in immunosuppressed patients who develop EBV lymphoma.